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Susan Wray

Publications and source records attributed to Susan Wray.

At least 37 records · Page 2Linked to original sources
PubMed2004

Cholecystokinin modulates migration of gonadotropin-releasing hormone-1 neurons.

Expression of the brain-gut peptide cholecystokinin (CCK) in the developing olfactory-gonadotropin-releasing hormone-1 (GnRH-1) neuroendocrine systems was characterized, and the function of CCK in these systems was analyzed both in vivo and in vitro. We present novel data demonstrating that CCK transcript and protein are expressed in sensory cells in the developing olfactory epithelium and vomeronasal organ, with both ligand and receptors (CCK-1R and CCK-2R) found on olfactory axons throughout prenatal development. In addition, migrating GnRH-1 neurons in nasal regions express CCK-1R but not CCK-2R receptors. The role of CCK in olfactory-GnRH-1 system development was evaluated using nasal explants, after assessing that the in vivo expression of both CCK and CCK receptors was mimicked in this in vitro model. Exogenous application of CCK (10(-7) m) reduced both olfactory axon outgrowth and migration of GnRH-1 cells. This inhibition was mediated by CCK-1R receptors. Moreover, CCK-1R but not CCK-2R antagonism caused a shift in the location of GnRH-1 neurons, increasing the distance that the cells migrated. GnRH-1 neuronal migration in mice carrying a genetic deletion of either CCK-1R or CCK-2R receptor genes was also analyzed. At embryonic day 14.5, the total number of GnRH-1 cells was identical in wild-type and mutant mice; however, the number of GnRH-1 neurons within forebrain was significantly greater in CCK-1R-/- embryos, consistent with an accelerated migratory process. These results indicate that CCK provides an inhibitory influence on GnRH-1 neuronal migration, contributing to the appropriate entrance of these neuroendocrine cells into the brain, and thus represent the first report of a developmental role for CCK.

Animals↗
PubMed2004

Electrophysiological characterization and functional importance of calcium-activated chloride channel in rat uterine myocytes.

In order to better understand the mechanisms underlying excitation of the uterus, we have elucidated the characteristics and functional importance of Ca(2+)-activated Cl(-) currents ( I(Cl-Ca)) in pregnant rat myometrium. In 101/320 freshly isolated myocytes, there was a slowly inactivating tail current (162+/-48 pA) upon repolarization following depolarising steps. This current has a reversal potential close to that for chloride, and was shifted when [Cl(-)] was altered. It was activated by Ca(2+) (but not Ba(2+)) entry through L-type Ca(2+) channels, enhanced by the Ca(2+) channel agonist Bay K8644 (2 microM), and inhibited by the Cl(-) channel blockers, niflumic acid (10 microM) and anthracene-9-carboxylic acid (9-AC, 100 microM). We therefore conclude that the pregnant rat myometrium contains Ca(2+)-activated Cl(-) channels producing inward current in ~30% of its cells. When these channels were inhibited by niflumic acid or 9-AC in intact tissues, the frequency of spontaneous contractions, was significantly reduced. Niflumic acid was also shown to inhibit oxytocin-induced contractions and Ca(2+) transients. Neither 9-AC nor niflumic acid had any effect on high-K-invoked contractions. Taken together these data suggest that Ca(2+)-activated Cl(-) channels are activated by Ca(2+) entry and play a functionally important role in myometrium, probably by contributing to membrane potential and firing frequency (pacemakers) in these cells.

Animals↗
PubMed2004

Functional architecture of the SR calcium store in uterine smooth muscle.

Sarcoplasmic reticulum (SR) is abundant in uterine smooth muscle cells. The functional role of this organelle in the regulation of uterine myocytes is not fully understood. The data available in the literature suggest that SR plays a dual role: as a source of calcium and as a calcium sink shaping calcium transients produced by membrane depolarisation and uterotonic agonists. Advances in digital imaging techniques including confocal microscopy of isolated living cells, and the development of methods for direct measurement of intraluminal calcium, has triggered a substantial increase in the number of publications elucidating the role of intracellular stores in calcium signalling. In this paper we review the literature and our own work on the SR calcium store in uterine smooth muscle cells.

Animals↗
PubMed2004

Dysfunctional labor and myometrial lactic acidosis.

OBJECTIVE: Inefficient uterine contractions are the most common cause of poor progress in labor. The global increase in cesarean delivery rate is a cause of considerable concern, and the greatest reason for increase is the result of failure to progress in labor. Following in vitro studies that showed acidification could depress uterine contraction, we hypothesized that it could contribute to dysfunctional labors. METHODS: A blood sample was taken from the lower segment of the uterus from women having a cesarean delivery, either electively or as a result of dysfunctional labor, and from those having a normal labor. This blood sample was analyzed for pH, O(2) saturation, and lactate levels. Contraction was recorded in myometrial strips, taken from women having elective cesarean delivery, at the pH of normally and dysfunctionally contracting uteri. RESULTS: The pH of myometrial capillary blood from women having a dysfunctional labor was significantly lower (7.35) than that from women having elective cesarean delivery (7.49) or cesarean delivery with normal contractions, with (7.47) or without (7.48) oxytocin (P <.001). The women in dysfunctional labor had higher capillary lactate and lower capillary O(2) saturation. Furthermore, in vitro, reducing the pH value from 7.5 to 7.3 changes regular uterine contractions to irregular ones of reduced amplitude. CONCLUSIONS: Myometrial lactic acidosis and a small decrease in O(2) saturation may be contributing factors to dysfunctional labor. Our data may also account for the ineffectiveness in management of dysfunctional labor with oxytocin. Oxytocin with a background of lactate acidosis may not be successful. LEVEL OF EVIDENCE: II-2

Acidosis, Lactic↗
PubMed2004

Ca2+ entry, efflux and release in smooth muscle.

Control of smooth muscle is vital for health. The major route to contraction is a rise in intracellular [Ca2+], determined by the entry and efflux of Ca2+ and release and re-uptake into the sarcoplasmic reticulum (SR). We review these processes in myometrium, to better understand excitation-contraction coupling and develop strategies for preventing problematic labours. The main mechanism of elevating [Ca2+] is voltage-gated L-type channels, due to pacemaker activity, which can be modulated by agonists. The rise of [Ca2+] produces Ca-calmodulin and activates MLCK. This phosphorylates myosin and force results. Without Ca2+ entry uterine contraction fails. The Na/Ca exchanger (NCX) and plasma membrane Ca-ATPase (PMCA) remove Ca2+, with contributions of 30% and 70% respectively. Studies with PMCA-4 knockout mice show that it contributes to reducing [Ca2+] and relaxation. The SR contributes to relaxation by vectorially releasing Ca2+ to the efflux pathways, and thereby increasing their rates. Agonists binding produces IP3 which can release Ca from the SR but inhibition of SR Ca2+ release increases contractions and Ca2+ transients. It is suggested that SR Ca2+ targets K+ channels on the surface membrane and thereby feedback to inhibit excitability and contraction.

Animals↗
PubMed2004

Mechanisms of action of pH-induced effects on vascular smooth muscle.

It is clear that pH has many effects on vascular smooth muscle and the overall action of pH on force will depend on the type of vascular smooth muscle in question and the combined effects on all the potential modulatory mechanisms. The major effects of pH on force appear to be mediated via modulation of [Ca]i rather than changes in the sensitivity of the contractile machinery to Ca2+. There are still numerous gaps in our understanding of the actions of pH and as more data become available, we will be able to better understand the major mechanisms involved.

Animals↗
PubMed2003

A new technique for simultaneous and in situ measurements of Ca2+ signals in arteriolar smooth muscle and endothelial cells.

We report here the first local and global Ca(2+) measurements made from in situ terminal arterioles. The advantages of the method are that there is minimal disturbance to the vessels, which retain their relationship to the tissue they are supplying (rat ureter) and the small size of vessel that can be studied. Good loading with the Ca(2+) indicator, Fluo-4 was obtained, and confocal sectioning through the tissue enabled vascular smooth muscle and endothelial cells to be clearly seen, along with red blood cells, nerve endings and the ureteric smooth muscle cells. We find the terminal arterioles to be extremely active, both spontaneously and in response to nor-adrenaline stimulation, with Ca(2+) sparks occurring in the vascular myocytes and Ca(2+) puffs in the endothelial cells. Even under resting conditions, endothelial cells produced oscillations and waves, which could pass from cell to cell, whereas the vascular myocytes only produced waves in response to agonist stimulation, and with no increase in the frequency of Ca(2+) sparks, and no spread from cell to cell. We compare our data to those obtained in dissected intact vessels and single cells. We conclude that this approach is a convenient and useful method for studying inter- and intracellular Ca(2+) signalling events and communication between cell types, particularly in very small vessels.

Action Potentials↗
PubMed2003

Calcium signaling and uterine contractility.

Changes in Ca(2+) signals within the myometrium have important functional consequences, as they determine contractility. We show that the basic phasic nature of uterine contractions, which is essential for successful labor, is critically dependent on Ca(2+) influx through voltage-gated L-type Ca(2+) channels, and hence in turn, on membrane potential. Thus changes in ion channel expression around term will play an important role in governing uterine excitability and contractility. There remains uncertainty about which channels are present in human myometrium and the nature of the pacemaker mechanism that initiates the action potential. The sarcoplasmic reticulum may augment, to a small extent, the necessary increase in [Ca(2+)] for contraction when agonists stimulate the uterus, but its main role appears to be to control excitability, acting as a negative feedback mechanism to limit contractions. Myosin light chain kinase activity and phosphorylation of myosin are essential components in the pathway of uterine contraction, once Ca(2+) has been elevated. Modulation of myosin light chain phosphatase activity can also influence contractions, but the effects are small compared with those modulating myosin light chain kinase. Ca(2+)-sensitizing pathways may not be utilized much in modulating normal phasic uterine activity, and caution is needed in extrapolating from in vitro experiments to in vivo conditions, especially because there may be redundant pathways. There is a need to study appropriate physiologic preparations, but these are not always available (eg, preterm laboring myometrium) and to combine functional studies with modern molecular approaches, to advance our understanding to a new level, from which better therapeutics will be developed.

Animals↗
PubMed2003

The effects of pH change on Ca(++) signaling and force in pregnant human myometrium.

OBJECTIVE: This study was designed to determine the effects of both intracellular and extracellular pH change on contractile activity and intracellular Ca(++) during spontaneous contractions, oxytocin, and depolarization-induced stimulation of human myometrium. STUDY DESIGN: Human myometrial tissue was obtained at elective caesarean delivery at term (37-41 completed weeks of gestation). Longitudinal strips were dissected and loaded with the calcium sensitive indicator Indo-1. Statistical significance was tested with the Student t test. RESULTS: Both intracellular and extracellular acidification significantly reduces or even abolishes phasic activity, whether it arises spontaneously or in the presence of oxytocin. These contractile changes can be accounted for by the changes in intracellular Ca(++). Alkalinization produced the opposite effects. However, baseline or maintained tension changes could not be accounted for by changes in intracellular Ca(++). CONCLUSION: We suggest that the effects on phasic activity are due to the inhibition of L-type calcium entry and that, during maintained or baseline activity, pH-sensitive Ca(++) release, possibly from the sarcoplasmic reticulum occurs; but it is insufficient to overcome the inhibitory effects at the myofilaments. We conclude that alterations of pH significantly affect calcium signaling and force production in the human myometrium and may contribute to dysfunction in labor.

Acids↗
PubMed2003

The effects of metabolic inhibition on intracellular calcium and contractility of human myometrium.

OBJECTIVE: Hypoxia occurs in the uterus during labour and may contribute to dysfunctional labours. We wanted to establish its effects on pregnant human myometrium and elucidate the mechanisms involved. DESIGN: Scientific study. SETTING: University Hospital and laboratories. POPULATION OR SAMPLE: Term pregnant women. METHODS: We measured contractions and intracellular [Ca(2+)] ([Ca(2+)](i)), in biopsies from term pregnant women undergoing elective caesarean section, and used cyanide to block oxidative phosphorylation. MAIN OUTCOME MEASURES: Changes in contractility and calcium. RESULTS: Although basal levels of [Ca(2+)](i) and tone rose, spontaneous and agonist-induced Ca(2+) transients and phasic contractions were rapidly reduced and abolished by cyanide. Neither stimulation of the uterus with oxytocin nor the Ca channel agonist, Bay K8644, prevented the changes produced by cyanide. The tonic force produced by depolarising the myometrium was also decreased by cyanide, but slowly recovered towards control levels, whereas [Ca(2+)](i) was maintained throughout. Similar data were obtained when nitrogen, rather than cyanide, was applied to the depolarised uterus. CONCLUSIONS: Impairment of oxidative phosphorylation is a potent depressor of phasic activity in human myometrium, irrespective of how it is produced, and our data suggest its effects lie at and beyond the surface membrane. Stimulation of the hypoxic uterus was not effective, which may explain the unpredictability of oxytocin application in some dysfunctional labours.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed2002

The role of the sarcoplasmic reticulum in neonatal uterine smooth muscle: enhanced role compared to adult rat.

Little is known about contractile activity, response to agonists or excitation-contraction coupling in neonatal smooth muscle. We have therefore investigated 10-day rat uterus to better understand these processes, and compared it to adult uterus to elucidate how control of contractility develops. Spontaneous contractions are present in the 10-day neonatal uterus, although they are not as large or as regular as those present in adult tissues. External Ca(2+) entry via L-type Ca(2+) channels is the sole source of Ca(2+) and is essential for the spontaneous activity. The neonatal uterus was responsive to carbachol or prostaglandin F(2alpha) application; it showed a marked stimulation and a clear dissociation between the force and Ca(2+) changes. Such sensitization was not apparent in adult rat myometrium. The sarcoplasmic reticulum (SR) had more releasable Ca(2+) and contributed more to the response to agonists in neonatal compared to adult tissues. Thus, Ca(2+) entry as opposed to SR Ca(2+) release contributed much less to the uterine response to agonists in the neonatal, compared to adult tissues. Inhibition of the SR by cyclopiazonic acid also caused a more vigorous increase in Ca(2+) and contractile activity, particularly frequency, in the neonatal compared to the adult uterus. Taken together these data suggest that: (1) spontaneous activity is already present by day 10, (2) receptor-coupling and excitation-contraction signalling pathways are functional, (3) the SR and Ca(2+) sensitization mechanisms play a more prominent role in the neonate, and (4) there is a shift to a greater reliance on Ca(2+) entry and excitability with development of the myometrium.

Aging↗
PubMed2002

In situ GABAergic modulation of synchronous gonadotropin releasing hormone-1 neuronal activity.

Evidence indicates that gonadotropin releasing hormone-1 [GnRH-1, also known as luteinizing hormone releasing hormone (LHRH)] neurons can exhibit synchronized neuroendocrine secretory activity before entrance into the CNS. In this study, we used calcium imaging to evaluate patterns of activity in individual, embryonic, GnRH-1 neurons as well as population dynamics of GnRH-1 neurons in mouse nasal explants maintained for 1 versus 3 weeks. Independent of age, GnRH-1 neurons displayed significant calcium peaks that synchronized at an interval of approximately 20 min across multiple GnRH-1 cells within an explant. Acute tetrodotoxin treatment decreased the amplitude of calcium peaks in individual GnRH-1 neurons and the duration but not the frequency of synchronized activity in the population of GnRH-1 neurons. Acute GABA(B) receptor antagonism increased the frequency of synchronized neuronal activity at both ages, whereas acute GABA(A) receptor antagonism decreased calcium oscillations in individual GNRH-1 cells as well as synchronization of the calcium pulses within the GnRH-1 population at the 1 week time point to background non-GNRH-1 cell levels. These results indicate that developing GnRH-1 neurons rely heavily on GABAergic signaling to initiate synchronized bouts of activity but thereafter, possess an innate capacity for synchronized activity patterns that are modulated by, but not completely dependent on GABAergic signaling.

Animals↗
PubMed2002

Characterisation of the ionic currents in freshly isolated rat ureter smooth muscle cells: evidence for species-dependent currents.

To better understand excitability, and hence contraction, the ionic currents underlying the action potential were identified and characterised in enzymatically isolated smooth muscle cells of the rat ureter. Using the whole-cell patch-clamp, under voltage-clamp conditions with K(+) in the pipette, three types of responses occurred to depolarisation: (1) sustained outward current and spontaneous transient outward currents (STOCs); (2) inward current; and (3) fast outward current. Investigation using different voltage protocols and pharmacological blockers and agonists revealed the presence of three outward and two inward currents. The outward currents were: (1) a sustained BK current, sensitive to low concentrations of tetraethylammonium (TEA) and featuring bursts of STOCs superimposed on it; (2) a fast, transient, A-type K current sensitive to 4-aminopyridine; and (3) a TEA and Ca(2+)-insensitive, late K(+) rectifier current. The inward currents were: (1) a fast L-type Ca(2+) channel current sensitive to nifedipine, Cd(2+) and potentiated by Ba(2+); and (2) a Ca(2+)-sensitive Cl(-) channel, which was inhibited by niflumic acid and Ba(2+), and produced a large tail current upon repolarisation at the end of the voltage step. The I- V relationships and peak amplitudes of all the currents are described. The finding of a K(+) rectifier and Ca(2+)-activated Cl(-) channel distinguish the rat ureteric cells from those of the guinea-pig. Thus, as well as the previously established difference in sarcoplasmic reticulum Ca(2+)-release mechanisms, there is also a species difference in ion channel expression in this tissue. We relate these currents to their possible contribution to the characteristically extremely long lasting action potential in the rat ureter.

4-Aminopyridine↗
PubMed2002

On the mechanisms whereby temperature affects excitation-contraction coupling in smooth muscle.

Moderate cooling of smooth muscle can modulate force production and may contribute to pathophysiological conditions, but the mechanisms underlying its effects are poorly understood. Interestingly, cooling increases force in rat ureter, but decreases it in guinea pigs. Therefore, this study used ureteric smooth muscle as a model system to elucidate the mechanisms of the effects of cooling on excitation-contraction coupling. Simultaneous recordings of force, intracellular [Ca(2+)], and electrical activity were made in intact ureter and ionic currents measured in isolated cells. The increase in force amplitude in rat ureter with cooling was found to be due to a significant increase in the duration of the Ca(2+) transient. This in turn was due to a marked prolongation of the action potential. In guinea pigs, both these parameters were much less affected by cooling. Examination of membrane currents revealed that differences in ion channel contribution to the action potential underlie these differences. In particular, cooling potentiated Ca(2+)-activated Cl(-) currents, which are present in rat but not guinea pig ureteric smooth muscle, and prolonged the plateau of the action potential and Ca(2+) entry. The force-Ca(2+) relationship revealed that the increased duration of the Ca(2+) transient was sufficient in the rat, but not in the guinea pig, to overcome kinetic lags produced in both species by cooling and potentiate force. Ca(2+) entry and release processes were largely temperature-insensitive, but the rate of relaxation was very temperature-sensitive. Effects of cooling on myosin light chain phosphatase, confirmed in experiments using calyculin A, appear to be the predominant mechanisms affecting relaxation. Thus, smooth muscle is diverse in its response to temperature, even when experimental variables, such as the mode of stimulation, are removed. Although the biochemical and mechanical events accompanying contraction are likely to be affected in similar ways by temperature, differences in electrical events lead to subsequent differences in these processes between smooth muscles.

Action Potentials↗
PubMed2002

Modulating signaling events in smooth muscle: cleavage of annexin 2 abolishes its binding to lipid rafts.

Cell membrane compartmentalization, which is believed to involve association of cholesterol- and glycosphingolipid-enriched membrane rafts, represents an important means of transmitting information across the plasma membrane. We have previously shown that raft association is mediated by the Ca2+-dependent binding of annexin 2 to the plasma membrane. In the present study, we demonstrate that the association of annexins 1 and 2 with the smooth muscle cell membrane can be terminated by their proteolytic cleavage. This proteolysis is thought to be triggered by calpain and occurs at non-raft regions of the plasma membrane. It is critically dependent on the intracellular concentration of free Ca2+ and requires an intact contractile apparatus. Annexins 1 and 2 interact with different membrane microcompartments--the former with non-raft, glycerolipid regions, the latter preferentially with membrane rafts. We demonstrate that PKC and RhoA, major signaling molecules that regulate smooth muscle contraction, are spatially segregated and interact with distinct membrane microcompartments. Proteolysis abolishes annexin binding to the plasma membrane and might result in rearrangement of membrane constituents followed by the interruption of segregation-dependent signaling events.

5'-Nucleotidase↗
PubMed2002

PH-induced changes in calcium: functional consequences and mechanisms of action in guinea pig portal vein.

The effects of changing extracellular (pH(o)) and intracellular pH (pH(i)) on force and the mechanisms involved in the guinea pig portal vein were investigated to better understand the control of tone in this vessel. When pH(o) was altered, the effects on force and calcium were the same irrespective of whether force had been produced spontaneously by high-K depolarization or by norepinephrine; alkalinization increased tone, and acidification reduced it. Because pH(o) changes also lead to changes in pH(i), we determined whether the effects on force could be explained by these induced pH(i) changes. It was found, however, that only with spontaneous activity did intracellular alkalinization increase force. In depolarized preparations, force was decreased, and, with norepinephrine, force was initially decreased and then increased. Thus the effects of pH(o) cannot be explained solely by changes in pH(i). The role of the sarcoplasmic reticulum (SR) and surface membrane Ca(2+)-ATPase on the mechanism were investigated and shown not to be involved. Therefore, it is concluded that both pH(o) and pH(i) can have powerful modulatory effects on portal vein tone, that these effects are not identical, and that they are likely to be due to effects of pH on ion channels rather than the SR or plasma membrane Ca(2+)-ATPase.

Ammonium Chloride↗